The invention is generally directed to systems and methods for high-throughput, real-time detection of analytes in fluids, for example, bodily fluids (e.g. cytokines in blood, glucose in saliva, tears and blood, etc.).
Current detection and screening techniques use low throughput, highly selective, non-scalable methods, which require labeling of the target molecule with a fluorophore to tag the specific molecule under study. These methods have several drawbacks: (1) prior knowledge of the molecule to be sensed is necessary, (2) modification of its structure is often needed to incorporate the tag, and (3) the tag molecule can change the way the primary molecule binds to other molecules, reducing the accuracy of an assay. Under alternative aspects of the present disclosure, label-free sensing is achieved using surface plasmon resonance (SPR) in thin metal films functionalized with specific linkers to selectively capture the analyte to be detected.
Typical SPR implementations rely on a prism or metallic grating (such as groove, slit and hole arrays) to couple the incident beam into propagating surface plasmon polaritons (SPPs), using light incident at a wavelength-specific angle. Also widely used are localized surface plasmon resonances (L-SPRs) in metal nanoparticles producing resonant scattering and extinction at specific frequencies. Given the resonant nature of the SPP excitation, practical implementations of SPR-based sensing schemes are limited in the number and range of wavelengths that can be used to sense the presence of analytes, thus further limiting their spectroscopic capabilities. Furthermore, SPR-based systems require large-volume samples and are limited to the detection of chemicals one at a time.
The dispersion relation of SPP waves shows longer wave vectors compared to electromagnetic waves traveling in dielectric materials (See FIG. 1a). This is why light incident upon a flat metal surface cannot excite surface plasmon polaritons directly (See FIG. 1b).
According to aspects of the present disclosure, excitation of SPP waves can be achieved by nano-corrugations, patterned on a flat metal surface (See FIGS. 1c-e), for example, grooves, slits, holes, or any surface plasmon launcher, which is any structure that can act as a SPP localized source.
FIG. 2a shows a commercially available Surface Plasmon Resonance (SPR) biochip. By measuring the angular shift of this characteristic band, detection of a refractive index change is accomplished. This approach leads to table-top instruments, where a large sensing area is needed to detect only one specific molecule per measurement session, thus limiting the throughput. Moreover, SPR sensors are monochromatic, in that they typically employ a single illumination wavelength, and they cannot determine the spectral fingerprints of the analyte.
Alternative approaches to label-free detection include fiber-optics, dielectric waveguides, nanowires, biochips, mechanical cantilevers, microring resonators, but none of the previous methods has shown significant throughput capabilities. For example, FIG. 2b shows an ultra high-Q microtoroid sensor.
Label-free detection can also be realized by measuring some optical properties of functionalized noble-metal nanoparticles, such as Surface Enhanced Raman Scattering (SERS), or Mie scattering and light extinction. For example, FIG. 2c shows a nanoscale biosensor. Unfortunately, the detection throughput is strongly limited by the difficulty to reliably address the response of several nanoparticles at once, by the challenging task to achieve uniform coating of all nanoparticles with a linker, and reproducible sensor response.
FIG. 2d shows a typical implementation of SPR, which involves a prism to excite the surface plasmons at the functionalized metal/dielectric interface, which happens when the incident angle is precisely chosen at a specified wavelength. In order to improve throughput, other techniques, such as SPR imaging, have been developed that allow several device cells (˜102) to be used in parallel to image the binding interaction and monitor intensity variations caused by a refractive index change in each cell.
Another method to excite SPR employs metallic gratings (such as hole, slit and groove arrays) as shown in FIGS. 2e-g. This method is based on the idea that the prism is replaced by a periodic array of scatterers etched in a metal film. Excitation of surface plasmons occurs at those resonant wavelengths (or angles, given a specific incident wavelength) satisfying the grating coupling condition. Therefore, reflection and transmission spectra from these devices are also characterized by sharp peaks corresponding to the wavelength- or angle-specific resonant condition. Metal nanoparticles are also widely used, which are characterized by sharp spectroscopic features in their scattering and extinction spectra, so-called localized surface plasmon resonances (LSPRs). Upon binding of the analyte of interest to the functionalized nanoparticle surface, a shift in the peak position of the LSPR is observed and the relative wavelength shift can be used to quantify the amount of analyte adsorbed at the surface. Given the “resonant” nature of the surface plasmon excitation, all the previous approaches involving SPR (prism-, grating-coupled, or localized) are limited in the number of wavelengths that can be used to sense the presence of the analyte. Therefore, the refractive index associated to the analyte can be measured only at one specific wavelength, thus strongly limiting the spectroscopic capabilities of any currently available SPR technique.
According to aspects of the present disclosure, a nanoscale plasmonic interferometer in one manifestation comprises of two grooves flanking a slit in a silver film is provided (see FIG. 2g).
A plasmonic-based device that accurately measures chemical and biological analytes in real-time is provided. Chemical analytes include but are not limited to dielectric materials such as semiconductor quantum dots (QDs) (see FIG. 3a), analytes embedded in thin films, or molecules in a gas or liquid phase. Biological analytes include but are not limited to proteins (e.g. cytokines in blood serum) and small molecules (e.g. glucose in bodily fluids) (see FIG. 3b-d).
In addition to a better understanding of light-matter interaction at the nanoscale, the disclosed methods and systems impact the throughput capabilities of several analyses and assays relevant to human health and currently used in the life sciences, and serve as an alternative high-throughput scheme for faster drug discovery, as well as more efficient identification and screening of novel therapies.